Rapid Acquisition of Multidimensional Solid-State NMR Spectra of Proteins Facilitated by Covalently Bound Paramagnetic Tags

Abstract: We describe a condensed data collection approach that facilitates rapid acquisition of multidimensional magic-angle spinning solid-state nuclear magnetic resonance (SSNMR) spectra of proteins by combining rapid sample spinning, optimized low-power radio frequency pulse schemes and covalently attached paramagnetic tags to enhance protein 1H spin-lattice relaxation. Using EDTA-Cu2+-modified K28C and N8C mutants of the B1 immunoglobulin binding domain of protein G as models, we demonstrate that high resolution and sensitivity 2D and 3D SSNMR chemical shift cor-relation spectra can be recorded in as little as several minutes and several hours, respectively, for samples containing ∼0.1-0.2 µmol of 13C,15N- or 2H,13C,15N-labeled protein. This mode of data acquisi-tion is naturally suited toward the structural SSNMR studies of paramagnetic proteins, for which the typical 1H longitudinal relaxation time constants are inherently a factor of at least ∼3-4 lower relativ